DESCRIPTION (the applicant's description verbatim): The goal of this competitive renewal application is to contribute to a mechanistic understanding of sudden death in heart failure. We will focus on alterations in gap junction communication in the heart which, we propose, occur as a fundamental response to myocyte injury and lead to the formation of anatomic substrates of arrhythmias. During the current grant interval, we made substantial progress in characterizing different expression patterns of multiple gap junction proteins (connexins) and identifying a tissue-specific spatial distribution of gap junctions in the normal heart that can account for distinct conduction properties of atrial and ventricular myocardium and specialized components of the conduction system. We also recently discovered that in the adult rat heart, the predominant ventricular connexin, Cx43, turns over with astonishingly rapidity (T1/2 = 1.3 hrs), indicating that even under basal conditions, gap junctions are highly dynamic structures. Using a variety of acute injury models in vitro and in vivo, we have observed striking changes in connexin +__ _____ __ ____ distribution and function that appear to be mediated by mechanisms involving connexin dephosphorylation and proteolysis. Accordingly, we now propose new studies designed to elucidate mechanisms of electrical uncoupling in response to acute myocyte injury and to characterize their acute and chronic electrophysiological consequences in the settings of ischemic heart disease and pressure-overload hypertrophy. To fulfill this goal, we will subject cultured myocytes to defined mechanical strain using a novel, highly versatile, in vitro stretch apparatus to elucidate the role of connexin dephosphorylation in proteolysis in the development of the conduction abnormalities analyzed with optical recordings of action-potential upstrokes. To apply insights gained in vitro to the intact heart subjected to pressure-overload and to gain further insights into potential roles of altered coupling in the development of arrhythmia substrates in decompensated ventricular hypertrophy, we will delineate mechanisms responsible for changes in connexin expression and gap junction structure and function in a widely used rat model of aortic constriction. We will also elucidate mechanisms responsible for electrical coupling in response to acute ischemia, and characterize the effects of changes in connexin phosphorylation, assembling gap junctions, and turnover dynamics in the pathogenesis of conduction disturbances during ischemia and reperfusion. The results of the proposed research will provide new insights into fundamental mechanisms of arrhythmogenesis in heart failure.